Nitrogenatd trans-stilbene analogs, method for the obtention and medical applications thereof

ABSTRACT

This invention is related to new nitrogenated trans-stilbene analog compounds, more specifically, imine, pyrrol and Indole derivatives, with procedures for the preparation and use thereof as pharmaceutical compositions for the treatment and/or chemoprevention of those mammalian diseases such as cancer, fibrosclerosis and acute/chronic inflammation, graft-versus-host reaction, ischemic-reperfusion tissue injury in stroke and heart attack, neurodegeneration, and during organ transplantation, whose pathogenic and pathophysiological mechanisms depend on or are significantly contributed by undesirable oxidative stress, angiogenic and proliferative responses.

This invention is related to new nitrogenated trans-stilbene analogcompounds, more specifically, imine, pyrrol and indole derivatives, withprocedures for the preparation and use thereof as pharmaceuticalcompositions for the treatment and/or chemoprevention of those mammaliandiseases such as cancer, fibrosclerosis and acute/chronic inflammation,graft-versus-host reaction, ischemic-reperfusion tissue injury in strokeand heart attack, neurodegeneration, and during organ transplantation,whose pathogenic and pathophysiological mechanisms depend on or aresignificantly contributed by undesirable oxidative stress, angiogenicand proliferative responses.

BACKGROUND OF THE INVENTION

It is known that the chemical synthesis of aromatic imines (or Schiffbases) can generally be achieved by means of the condensation betweenprimary amines and carbonylic compounds (cf. Patai, The Chemistry ofCarbon-Nitrogen Double Bond; Wiley: New York, 1970, pp. 64).

Likewise, polysubstituted pyrrol rings can be chemically synthesized indifferent ways by employing linear or convergent synthesis methodologies(cf. Sundberg, Comprehensive Heterocyclic Chemistry; Katrizki, A. andRees, C. W. Eds.; Pergamon: Oxford, 1984; vol. 4, pp. 313). Onesufficiently general way consists of the aromatization of substitutedpyrrolidines (cf. Fejes et al. Tetrahedron 2000, vol. 56, pp. 8545;Gupta et al. Synth. Commun. 1998, vol. 28, pp. 3151). The finalheterocycles can be prepared, in turn, by means of the 1,3-dipolarreaction between azomethine ylides and electronically deficient alkenes(cf. Ayerbe et al. J. Org. Chem. 1998, vol. 63, pp. 1795; Vivanco et al.J. Am. Chem. Soc. 2000, vol. 122, pp. 6078).

Methods also abound for the synthesis of indoles (cf. Horton et al.Chem. Rev. 2003, vol. 103, pp. 893), one of which is a convergentprocedure described in literature consisting of the thermal cyclationbetween primary arylamines and haloacetopheonone derivatives (cf.Nyerges et al. Tetrahedron Lett. 2005, vol. 46, pp. 377). However, theyields obtained by this method are not usually very high, due mainly tothe relatively high temperatures and the lengthy reaction timesnecessary for completing the cyclation reaction and the ease with whichthe reagents may degrade under these conditions.

Oxidative stress facilitates carcinogenesis (Engel R H, Evens A M.Oxidative stress and apoptosis: a new treatment paradigm in cancer.Front Biosci. 2006; 11:300-12) and the prometastatic and proangiogenicmechanisms of cancer (Tanaka T, Akatsuka S, Ozeki M, Shirase T, Hiai H,Toyokuni S. Redox regulation of annexin 2 and its implications foroxidative stress-induced renal carcinogenesis and metastasis. Oncogene.2004; 23:3980-9) and many other diseases (Casetta I, Govoni V, GranieriE. stress, antioxidants and neurodegenerative diseases. Curr Pharm Des.2005; 11(16):2033-52; Sukkar S G, Rossi E. Oxidative stress andnutritional prevention in autoimmune rheumatic diseases. Autoimmun Rev.2004; 3:199-206; Naito Y, Takano H, Yoshikawa T. Oxidativestress-related molecules as a therapeutic target for inflammatory andallergic diseases. Curr Drug Targets Inflamm Allergy. 2005; 4:511-5.).Thus, numerous cancer research studies have focused their attention onthe effects of some natural antioxidant compounds (resveratrol,quercetin, vitamin C, etc.) as chemopreventive agents of carcinogenesisand metastasis. In come cases, it has been proven that said action iscaused not solely by means of the antioxidant effect of said agents, butalso by way of their action of blocking cyclooxygenases (COX) andtyrosine kinases.

On the other hand, the mechanism of action of most of theproinflammatory and prometastatic factors is regulated byoxygen-reactive metabolites. In this regard, it has been proven thattreating animals with catalase prior to their being intrasplenicallyinoculated with B16 melanoma cells reduces the onset of hepaticmetastasis, which indicates that hydrogen peroxide (H2O2), released inresponse to hepatic colonization by tumor cells, has prometastaticeffects (cf. Anasagasti et al., “Sinusoidal endothelium release ofhydrogen peroxide enhances very late antigen-4-mediated melanoma celladherence and tumor cytotoxicity during interleukin-1 promotion ofhepatic melanoma metastasis in mice”. Hepatology. 1997, vol. 25 pp.840-6).

Trans-stilbene compounds, particularly trans-resveratrol, are widespreadthroughout nature, mainly in the form of phytoalexins and are attractinggrowing interest due to a wide range of biological activities useful inoncology, such as the inhibition of carcinogenesis (cf. Jang et al.,Science 1997, vol. 275, pp. 218; Gosslan et al., Brit. J. Cancer. 2005,vol. 92, pp. 513) and apoptosis induction (cf. Lee et al., Life Sci.2004, vol. 75, pp. 2829). This biological activity has been attributedto the antioxidant properties (cf. Stivala et al., J. Biol. Chem. 2001,vol. 276, pp. 22586) and anti-inflammatory properties (cf. Kimura etal., Biochim. Biophys. Acta 1985, vol. 834, pp. 275) of these compounds,as a result of which they can serve as chemopreventive andchemotherapeutic agents (cf. De Lédinghen et al., Int. J. Oncol. 2001,vol. 19, pp. 83; Scheneider et al., Nutr. Cancer 2001, vol. 39, pp. 102;Mahyer-Roemer et al., Int. J. Cancer 2001, vol. 94, pp. 615). It isknown that trans-stilbenes are able to isomerize the cis-form which iseither inactive or less active. For example, trans-resveratrol canisomerize under the effect of sunlight to convert partially into the cisisomer (cf. F. Olalla, Curr. Med. Chem. 2006, vol 13, pp. 87-98; 1.Kolouchová-Hanzliková et al. Food Chem. 2004, vol. 87, pp. 151-158).

DESCRIPTION OF THE INVENTION Brief Description of the Invention

The invention described herein is related to nitrogenatedpolyhydroxylated and/or polyalcoxylated trans-stilbene analog compounds,which include imine derivatives, pyrrol derivatives or indolederivatives, useful as inhibitors of inflammatory agents, oxidativestress, angiogenesis-related effects, metastasis and cancer progression.This invention is also related to therapeutic compositions which includesaid compounds and the use thereof for the treatment and prophylaxis ofcancerous and inflammatory diseases, such as, but not limited to, cancermetastasis. This invention is also related to the methods for obtainingsaid compounds.

DESCRIPTION OF THE FIGURES

FIG. 1.—Shows the optimized structures, calculated using the MP3 method(cf. Stewart, J. Comput. Chem. 1989, vol. 10 pp. 209), of (A)trans-stilbene; (B) diphenylmine; (C) 2,4-diphenyl-1H-pyrol; y (D) 2phenyl-1H-indole.

FIG. 2.—Shows the inhibitory effect of the compounds JE1:2, JE2:1 andJE2:2 on the adhesion of the murine B16 melanoma (MB16) cells treated ornot with non-toxic concentrations of H2O2 to immobilized recombinantVCAM-1 substrates. The differences in adhesion with regard to theuntreated MB16 cells (*) and the MB16 cells treated with H2O2 (**) arestatistically significant (P<0.01) according to Student's t-test.

FIG. 3.—Shows the inhibitory effect of the compounds JE1:2, JE2:1 andJE2:2 on the in vitro proliferation of MB16 cells treated or not withrecombinant IL-18. The differences in proliferation with regard to theuntreated MB16 cells (*) and the MB16 cells treated with IL-18 (**) arestatistically significant (P<0.01) according to Student's t-test.

FIG. 4.—Shows the inhibitory effect of the compound JE2:2 on theproduction of H2O2 from MB16, treated or not with recombinant IL-18 invitro.

FIG. 5.—Shows the inhibitory effect of the compounds JE1:2, JE2:1 andJE2:2 on the production of H2O2 from primary cultured mouse hepaticsinusoidal endothelium (HSE) cells.

FIG. 6.—Shows the inhibitory effect of the compound JE2:2 on theadhesion of MB16 cells to the primary cultured mouse HSE cells treatedor not with MB16 conditioned media (MC-MB16). The differences inadhesion with regard to the untreated HSE cells (*) and the HSE cellstreated with MC-MB16 (**) are statistically significant (P<0.01)according to Student's t-test.

FIG. 7.—Shows the effects of the compounds JE2:2-01 and JE2:1-02 on theadhesion of the MB16 cells treated or not with H2O2 to immobilizedVCAM-1 substrates. The differences in adhesion with regard to theuntreated MB16 cells (*) and the MB16 cells treated with H2O2 (**) arestatistically significant (P<0.01) according to Student's t-test.

FIG. 8.—Shows the inhibitory effect of JE2:2-01 and JE2:1-02 (A) on theadhesion of MB16 cells and (B) on the production of TNF-alpha from HSEcells treated or not with MC-MB16. The differences in the adhesion or inthe concentration of TNF-alpha with regard to the untreated HSE cells(*) and the HSE cells treated with MC-MB16 (**) are statisticallysignificant (P<0.01) according to Student's t-test.

FIG. 9.—Shows the inhibitory effect of the compounds YEF02, YEF03,YEF07, YEF05B, YEF07B and YEF05H on the adhesion of MB16 cells treatedor not with H2O2 to immobilized VCAM-1 substrate. The differences inadhesion between untreated MB16 cells (*) and H2O2-treated MB16 cells(**) are statistically significant (P<0.01) according to Student'st-test.

FIG. 10.—Shows the inhibitory effect of the compounds YEF02, YEF03,YEF07, YEF05B, YEF07B and YEF05H on the adhesion of MB16 cells to HSEtreated or not with MC-MB16. The differences in adhesion with regard tountreated (*) and the MC-MB16-treated (**) HSE cells are statisticallysignificant (P<0.01) according to Student's t-test.

FIG. 11.—Shows the inhibitory effect of the compounds JE22, YEF07 andYEF05B on the production of PGE2 (as evidence of cyclooxygenase-2activity) by HSE cells in response to VEGF. The differences in the PGE2production between untreated (*) and VEGF-treated (**) HSE cells arestatistically significant (P<0.01) according to Student's t-test.

FIG. 12A, FIG. 12B y FIG. 12C.—Effect of the compound JE2:2 on thedevelopment of hepatic metastasis following the intrasplenic injectionof basal medium-cultured MB16 cells in C57BL/6J mice.

DETAILED DESCRIPTION OF THE INVENTION

According to the first aspect of the invention described herein, anitrogenated compound of polyhydroxylated and/or polyaloxylatedtrans-stilbene is provided with the following general formula (I):

or any of the salts thereof, where:(X) is selected from between the following groups, imine or pyrrole:

In the case in which the group (X) is a pyrrole group, it may be bondedto the phenyl group (A) by one or two of its pyrrole ring carbons. Whenit is bonded by two of its carbons, then (D) and (Y) form part of theortho-disubstituted phenyl group (A);

(D) is the phenol ring (A) (aromatic ring (A) of the general structure(I);(U) may be selected from among a hydrogen atom, a linear or branchedalkyl group (C₁-C₁₀) or the phenyl group (B) (aromatic ring (B) ofgeneral formula (I);(Y) may be selected from among a hydrogen atom or a group selected fromamong nitro (NO₂), amino (NR₂), linear or branched alkoxycarbonyl(—C(═O)OR), amide (NRC(═O)R′) or an organic or inorganic quaternaryammonium salt (NR₄ ⁺), such as, for example but not limited to,quaternary ammonium chloride or tartrate;(Y) does not exist when the carbon to which it is bonded forms part ofthe phenyl A ortho-disubstituted group(W) may be selected from among a hydrogen atom or a group selected amongcarboxyl (—C(═O)OR) or aminocarbonyl [(C(═O)NRR′) mono or disubstitutedfor alkyl, aryl or heteroaryl groups] oe (W) is the phenyl group (B)(aromatic ring (B) of general formula (I)(Z) may be selected from among a hydrogen atom or a group selected fromamong a linear or branched alkyl (C₁-C₁₀), benzyl (—C₆H₅), carboxyl andanalogs (—C(═O)OR), arylakyl, heteroarylmethyl, O-alkyl(aryl)carbamoylor N-alkyl(aryl)semicarbazide.

R¹, R², R³, R⁴, R⁵, R⁶, R⁷, R⁸, R⁹ and R¹⁰ may be the same or differentand are selected from among a hydrogen atom or a group selected betweenalkoxy (—OR) and hydroxyl (—OH), where at least three of thesesubstitutes are either alkoxy and/or hydroxyl.

R⁵ is absent when the pyrrole ring is bonded to ring (A) by two of itscarbon atoms.

In the invention described herein, the term “alkyl” C₁-C₁₀, is asaturated linear or branched-chain hydrocarbon which includes from 1 to10 carbon atoms. The alkyl groups preferred in this invention are, butare not limited to, those which have 1 to 5 carbon atoms, for example,the methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl,tert-butyl, pentyl and isopentyl groups.

The term “alkoxy” means an (—OR) radial, where R is a linear or branchedalkyl chain of 1 to 10 carbon atoms. The alkoxy groups preferred in thisinvention are, but are not limited to, those having 1 to 5 carbon atoms,such as, for example, methoxyl, etoxyl, propoxyl, isopropoxyl, butoxyl,pentoxyl, sec-butoxyl or tert-butoxyl.

The term “amine” means a radical (—NR₂), where the two R groups may bethe same or different and represent a hydrogen atom or a linear orbranched alkyl chain of 1 to 10 carbon atoms. The amino groups preferredin this invention are, but are not limited to, those in which both R arehydrogen of any thereof is an alkyl chain of 1 to 5 carbon atoms, morepreferably of 1 or 2 carbon atoms.

The term “arylalkyl” means a linear or branched chain of 1 to 5 carbonatoms which is substituted by an aryl radical, where the term “aryl”, inthis invention, means a substituted or non-substituted phenyl radical.The group is preferably, but not limited to, an arylmethyl.

The term “heteroarylalkyl” means a linear or branched chain or 1 to 5carbon atoms which is substituted by an aromatic radical of 5 or 6 bondswith one, two or three heteroatoms, understood as being the elementsnitrogen, oxygen and sulfur.

The term “carboxyl” (—C(═O)OR) encompasses a carboxylic acid (R═H) andan ester in which R may be a linear or branched alkyl group (linear orbranched “alkoxycarbonyl”), or a cyclic alkyl group.

The term “amide” means a radical with the form (—NCR(═O)R′) where the Rand R′ groups may be the same or different and represent an atom ofhydrogen or a linear or branched alkyl chain of 1 to 10 carbon atoms.The amide groups preferred in this invention are, but are not limitedto, those in which both R's are hydrogen and one thereof is an alkylchain of 1 to 5 carbon atoms, more preferably of 1 or 2 carbon atoms.

The term “aminocarbonyl” means a radical with the form (—C(═O)NRR′)where the R and R′ groups may be the same or different and represent ahydrogen atom or a linear or branched alkyl chain of 1 to 10 carbonatoms or an aryl or a heteroaryl group.

In one preferred embodiment of the invention described herein, a formula(II) compound is provided, which is obtained on substituting (X) in thegeneral formula (I) with an imine group:

or any of the salts thereof, where:R¹, R², R³, R⁴, R⁵, R⁶, R⁷, R⁸, R⁹ and R¹⁰ are of the same meaning aspreviously stated hereinabove for the general formula (I).

In a more preferred embodiment of the invention described herein, thegeneral formula (II) compound is, but is not limited to,5-((E)-(4-Hydroxyphenylimine)methyl)benzene-1,3-diol.

In another preferred embodiment of the invention described herein, acompound with the formula (III) is provided, which is obtained onsubstituting (X) in the general formula (I) with a pyrrole group:

or any of the salts thereof, whereR¹, R². R³, R⁴, R⁵, R⁶, R⁷, R⁸, R⁹, R¹⁰ and group (Z) are of the samemeaning as previously stated hereinabove for the general formula (I);(Y) may be selected from among a hydrogen atom or a group selected fromamong nitro (NO₂), amine (NR₂), linear or branched alkoxycarbonyl(—C(═O)OR), amide (—C(═O)N—RR′) or aminocarbonyl NRC(═O)R′) or anorganic or inorganic quaternary ammonium salt (NR₄ ⁺), such as, forexample, but not limited to, quaternary ammonium chloride or tartrate;(W) may be selected from among a hydrogen atom or a group selectedbetween carboxyl (—C(═O)OR) or aminocarboxyl (—C(═O)NRR′).

One preferred embodiment of the invention described herein entails ageneral formula (III) compound where (Z) is a hydrogen atom; (W) is agroup selected between —COOCH₃ or —COOH; (Y) is a hydrogen atom or agroup selected between —NH₂ or —NO₂; R¹, R⁵, R⁶ are R¹⁰ a hydrogen atom;and R², R³, R⁴, R⁷, R⁸ and R⁹ are the same or different and represent ahydrogen atom or a group selected between —OCH₃ or —OH.

A more preferred embodiment of the invention described herein entails ageneral formula (III) compound where (Z) and (Y) are hydrogen atoms.

An even more preferred embodiment of the invention described hereinentails a general formula (III) compound selected, but withoutlimitation, from among the following group:3-(4-methoxyphenyl)-5-(dimethoxyphenyl)-1H-pyrol-2-carboxyilic acid,3,5-bis(3,5-Dihydroxyphenyl)-1H-pyrol-2-methyl carboxylate,3,5-bis(3,5-Dimethoxyphenyl)-1H-pyrol-2-methyl carboxylate,5-(3,5-Dihydroxyphenyl)-3-(4-hydroxyphenyl)-1H-pyrol-2-methylcarboxylate,5-(3,5-dimethoxyphenyl)-3-(4-methoxyphenyl)-1H-pyrol-2-methylcarboxylate,5-(3,5-dimethoxyphenyl)-3-(4-methoxyphenyl)-1H-pyrol-2-methylcarboxylate,5-(3,5-dimethoxyphenyl)-3-(4-methoxyphenyl)-4-nitro-1H-pyrol-2-carboxylate,4-amino-3,5-bis(3,5-dimethoxyphenyl)-1H-pyrol-2-methyl carboxylate,5-(3,5-dimethoxyphenyl)-3-(4-methoxyphenyl)-4-nitro-1H-pyrol-2-methylcarboxylate, 3,5-bis(3,5-dimethoxyphenyl)-4-nitro-1H-pyrol-2-methylcarboxylate, 3,5-bis(3,5-dimethoxyphenyl)-1H-pyrol-2-carboxylic acid,and 3-(3,5-Dihydroxyphenyl)-5-(4-hydroxyphenyl)-1H-pyrol-2-methylcarboxylate.

One further embodiment of the invention described herein provides aformula (VI) compound which is obtained on substituting (X) in thegeneral formula (I) with a pyrrole group bonded to the aromatic ring (A)by two of its carbon atoms:

or any of the salts thereof, whereR¹, R², R³, R⁴, R⁶, R⁷, R⁸, R⁹ and R¹⁰ and the group (Z) are of the samemeaning as previously stated hereinabove for general formula (I); and(U) is hydrogen or a linear or branched C₁-C₁₀ alkyl group

A preferred embodiment of the invention described herein entails ageneral formula (IV) compound in which (Z) is a hydrogen atom or a—COOC(CH₃)₃ group; (U) is a hydrogen atom; and R¹, R², R³, R⁴, R⁶, R⁷,R⁸, R⁹ and R¹⁰ are the same or different and represent a hydrogen atomor a group selected between —OCH₃ or —OH.

A more preferred embodiment of the invention described herein entails ageneral formula (IV) compound selected, but without limitation, fromamong the following group: 2-(2,5-dihydroxyphenyl)-1H-indole-4,6-diol;4,6-Dimethoxy-2-(3,5-dimethoxyphenyl)-1H-indole;4,6-Dimethoxy-2-(2,5-dimethoxyphenyl)-1H-indole-1-tert-butylcarboxylate; 4,6-Dimethoxy-2-(2,5-dimethoxyphenyl)-1H-indole; and4,6-Dimethoxy-2-(3,5-dimethoxyphenyl)-1H-indole-1-tert-butylcarboxylate.

The term “analogs” means, in this description, compounds of a similarstructure, in other words, is bearing a similarity on the periphera ofsaid molecules.

As is shown in FIG. 1, in the substituted trans-stilbenes, the distancesbetween the quaternary aromatic atoms which bond the phenyl groups tothe central group are on the order of 3.82 Å, a value within a mid-rangeas compared to that calculated for N-benzylidene anilines I (ca. 3.74 Å,FIG. 1B), 2,4-diaryl-1H-pyrroles (ca. 5.06 Å, FIG. 1C) and2-aryl-1H-indoles (each 3.71 Å, FIG. 1D). Likewise, some hypotheticalhydroxyl or alkoxy groups in a relative arrangement similar to that ofthe trans-resveratrol are spaced at the intervals of 10.16-8.78 Å and9.39-7.73 Å for compounds I, II and III, values which encompass those ofthe resveratrol proper (8.85 Å and 8.55 Å respectively, FIG. 1A).Therefore, the compounds of the invention make it possible to extend thetherapeutical applications of the Trans-stilbenes on broadening thegeometric parameters of the pharmacophore and to provide the possibilityof optimizing interactions of major importance for the desiredpharmacological activity.

The compounds described are of the following structural formula:

5-((E)-(4-Hydroxyphenylimino)methyl)benzene-1,3-diol, referred tohereinafter as “Azaresveratrol”:

3,5-bis(3,5-Dihydroxyphenyl)-1H-pyrrole-2-methyl carboxylate, referredto hereinafter as “JE2:2”:

3,5-bis(3,5-Dimethoxyphenyl)-1H-pyrrole-2-methyl carboxylate, referredto hereinafter as “JEM2:2-01”:

5-(3,5-Dihydroxyphenyl)-3-(4-hydroxyphenyl)-1H-pyrrole-2-methylcarboxylate, referred to hereinafter as “JE2:1”:

5-(3,5-dimethoxyphenyl)-3-(4-methoxyphenyl)-1H-pyrrole-2-methylcarboxylate:

3-(3,5-Dihydroxyphenyl)-5-(4-hydroxyphenyl)-1H-pyrrole-2-methylcarboxylate, referred to hereinafter as “JE1:2”:

5-(3,5-dimethoxyphenyl)-3-(4-methoxyphenyl)-1H-pyrrole-2-methylcarboxylate:

3,5 acid 3,5-bis(3,5-dimethoxyphenyl)-1H-pyrrole-2-carboxylic acid:

3-(4-methoxyphenyl)-5-(dimethoxyphenyl)-1H-pyrrole-2-carboxylic acid,referred to hereinafter as “JEM2:1-02”:

5-(3,5-dimethoxyphenyl)-3-(4-methoxyphenyl)-4-nitro-1H-pyrrole-2-carboxylate:

3,5 acid 3,5-bis(3,5-dimethoxyphenyl)-1H-pyrrole-2-methyl carboxylate:

5-(3,5-dimethoxyphenyl)-3-(4-methoxyphenyl)-4-nitro-1H-pyrrole-2-methylcarboxylate:

3,5-bis(3,5-dimethoxyphenyl)-4-nitro-1H-pyrrole-2methyl carboxylate:

2-(2,5-Dihydroxyphenyl)-1H-indole-4,6-diol, referred to hereinafter as“YEF05H”:

4,6-Dimethoxy-2-(3,5-dimethoxyphenyl)-1H-indole, referred to hereinafteras “YEF07”:

4,6-Dimethoxy-2-(2,5-dimethoxyphenyl)-1H-indole:

4,6-Dimethoxy-2-(2,5-dimethoxyphenyl)-1H-indole-1-tert-butylcarboxylate, referred to hereinafter as “YEF07B”:

4,6-Dimethoxy-2-(3,5-dimethoxyphenyl)-1H-indole-1-tert-butylcarboxylate, referred to hereinafter as “YEF05B”:

A second aspect of the invention described herein is the method forobtaining the general formula (II) compound, which entails reacting inthe presence or absence of an organic solvent:

-   -   i) an aromatic aldehyde suitably substituted for alkoxy or        hydroxyl groups of the following formula (V),

wherein:R¹, R², R³, R⁴ and R⁵ are of the same meaning as previously statedhereinabove;

-   -   ii) an aniline of formula (VI) similarly substituted

wherein: R⁶, R⁷, R⁸, R⁹ and R¹⁰ are of the same meaning as previouslystated hereinabove.

A preferred embodiment of the method for obtaining the formula (II)compound employs a drying agent selected, without limitation, betweeneither a suitable anhydrous salt or molecular sieve.

After eliminating the solvent and the drying agent, in the event thateither one thereof is used, the resulting reaction mixture is purifiedby means of crystallization in the suitable solvent known by any experton the subject.

A third aspect of the invention described herein provides a method forobtaining the formula (III) compound which entails reacting:

-   -   i) an (E) or (Z)-configuration imine of the following formula        (VII):

wherein: R¹, R², R³, R⁴ and R⁵ are of the same meaning as previouslystated hereinabove, and R¹¹ represents an alkyl (C₁-C₁₀), preferablymethyl, an aryl or an heteroaryl group;

-   -   ii) an (E) or (Z)-configuration nitroalkene of the formula        (VIII):

where: R⁶, R⁷, R⁸, R⁹ and R¹⁰ are of the same meaning as previouslystated hereinabove;

-   -   iii) a metallic salt, selected, without limitation, from among        lithium perchlorate, silver perchlorate or silver acetate; and    -   iv) a tertiary organic base selected from among the aliphatic        bases with C₃-C₁₀ carbons or alkane-aromatic bases with C₉-C₁₅        carbons.

The reaction mixture can be made by means of microwave radiation or byadding one of the components to the other three, in an organic solventand at the temperature of −25° C. to +25° C., preferably at temperaturesnearing +25° C.

The completion of the cycloaddition reaction results in obtaining amixture of 2-alkoxycarbonil pyrroleidines corresponding to thesubstitutes selected for each individual reaction. Said mixture isdissolved in a cyclic ether such as high-boiling-point acyclic ortetra-hydrofurane such as diethylene glycol dimethyl ether, also knownas “diglyme” and an oxidizing agent added such as manganese dioxide,hydrogen peroxide or 2,3-dichloride-5,6-dicyano-1,4-benzoquinone. Aftera certain length of time at temperatures ranging from +60° C. to +250°C., the corresponding mixture is obtained comprised of2-alkoxycarbonyl-NH-pyrrole (IX) and 2-alkoxycarbonyl-4-nitro-NH-pyrrole(X) the components of which can be separated by means of fractionedcrystallization or chromatography:

These (IX and (X) compounds pertain to the family of compounds ofgeneral formula (III) included in the invention described herein, in theparticular cases in which (Y) is NO₂ or H; (W) is COOR¹¹; (Z) is H andR¹-R¹⁰ may be the same or different and are selected from among ahydrogen atom or a group selected between either alkoxy (—OR) orhydroxyl (—OH), where at least three of these substitutes are eitheralkoxy and/or hydroxyl;

For all of the other cases, the starting compounds are (IX) and (X), andthe following chemical transformations are carried out:

To obtain the general formula (III) compounds, wherein (Y) is NH₂ andthe derivatives thereof, the nitro group is reduced to the amine grouppreferably by reacting the corresponding nitrocompound with tindichloride at temperatures ranging from +25° C. to +90° C. Following thepurification of the corresponding primary amine, this amine may betransformed into derivatives of the substituted amine, amide or ammoniumsalt type by means of conventional processes.

To obtain the formula (III) compounds wherein (W) is COOH, thehydrolysis of the ester function present in the compounds (IX) and (X)is preferably achieved by means of the treatment thereof with sodium orlithium hydroxide in a mixture of water and dimethoxyethane.

To obtain the formula (III) compounds wherein (W) is H, thedecarboxylation of the carboxylic acids obtained in the precedingparagraph hereinabove is preferably achieved by means of thermolysis atpressures ranging from 5 mm Hg to 0.01 mm Hg and at temperatures rangingfrom +150° C. and +250° C.

To obtain the compounds of formula (III) wherein R¹-R¹⁰ are OH, thetransformation of the methoxyl group into hydroxyl groups is preferablyachieved by reacting the corresponding methoxylate compound under aninert atmosphere preferably of argon, with a solution of borontribromide in dichloromethane at temperatures ranging from −20° C. to+30° C. for 2-24 hours, following which the mixture is treated withmethanol at 0° C.

To obtain the formula (III) compound where (Z) may be alkyl (C₁-C₁₀),aryl, O-alkyl(aryl)carbamoyl or N-alkyl(aryl)-semicarbazide. Thesubstitution of the carbon atom present in the (IX) and (X) compounds isachieved by means of conventional alkylation and acylation methods. Inthe cases in which inertia is observed in the acylation, this may befacilitated by means of the use of anhydrides a catalyst acylatingagents, preferably 4-dimethylaminopyridine and zinc perchlorate.

A fourth aspect of the invention described herein provides a method forobtaining the formula (VI) compound, which entails reacting:

-   -   i) an amine of formula (XI) below,

wherein: R¹, R², R³ and R⁴ are of the same meaning as previously statedhereinabove;

-   -   ii) an alpha-halocetone of formula (XII) below,

wherein: (G) may be a halogen selected from chlorine, bromine andiodine;(U) is of the same meaning as previously stated hereinabove, andR⁶, R⁷, R⁸, R⁹ and R¹⁰ are of the same meaning as previously statedhereinabove;

-   -   iii) a tertiary amine such as N,N-dimethylaniline or any        combination of aryl, heteroaryl or linear, cyclic or branched        alkyl groups.

For the purposes of the invention described herein, the reaction mixturecomprised of the three immediately preceding components statedhereinabove can be achieved by means of microwave radiation in theabsence of a solvent at a temperature ranging from +90° C. to +180° C.,preferably at temperatures nearing +150° C., with a radiation power of25 to 200 W, preferably employing radiation power nearing 100 W, forradiation times of 5 to 30 minutes, the preferred radiation times beingof around 10 minutes. The radiation may be taken to atmospheric pressureor to pressures of 50 to 200 psi (Pounds/square inch). Upon completionof the reaction, general formula (XIII) 2-aryl-1H-indoles are obtained.

Said (XIII) compounds pertain to the general formula (IV) compoundsincluded in the invention described herein, in the particular cases inwhich (Z) is H and R¹-R¹⁰ are of the same meaning as previously statedhereinabove.

For all of the other cases, starting from the (XIII) compounds, thefollowing transformations are carried out:

To obtain the formula (IV) compound wherein any of the R¹-R¹⁰ groups areOH, the transformation of the methoxyl groups into hydroxyl groups ispreferably achieved by reacting the corresponding methoxylated compoundunder an inert atmosphere preferably of argon, with a solution of borontribromide in dichloromethane at temperatures ranging from −20° C. to+30° C. for 2-24 hours, following which the mixture is treated withmethanol at 0° C.

To obtain the formula (IV) compound wherein (Z) can be alkyl (C₁-C₁₀),aryl, O-alkyl(aryl)carbamoyl or N-alkyl(aryl)-semicarbazide: thesubstitution of the hydrogen atom present in the XIII compounds isachieved by means of conventional alkylation and acylation methods. Inthose cases in which inertia is observed in the acylation, the acylationcan be facilitated by means of employing anhydrides as catalystacylating agents, preferably 4-dimethylaminopyridine and zincperchlorate.

A fifth aspect of the present invention provides the use of the generalformula (I) compounds for the treatment or chemoprevention of thosemammalian diseases such as cancer, fibrosclerosis and acute/chronicinflammation, graft-versus-host reaction, ischemic-reperfusion tissueinjury in stroke and heart attack, neurodegeneration, and during organtransplantation, whose pathogenic and pathophysiological mechanismsdepend on or are significantly contributed by undesirable oxidativestress, angiogenic and proliferative responses.

A sixth aspect of the invention provides the use of any of the generalformula (I) compounds or combinations thereof for the preparation of apharmaceutically-acceptable composition for the treatment andprophylaxis of diseases involving cancerigenous and inflammatoryprocesses, more preferably, but without limitation to, the treatment ofhepatic metastasis.

One embodiment of the preparation of a composition may be, without beinglimited to, that of a composition which includes at least one of thegeneral formula (II) compounds and one or morepharmaceutically-acceptable excipients. A composition may likewise beprepared for the formula (III) and (IV) compounds. The formula (II),(III) and (IV) compounds of the invention described herein may beadministered both as a pure substance as well as in the form ofpharmaceutical formulations, although the administration of thecombined-form composition is preferable.

Another aspect of the invention described herein provides apharmaceutical composition which includes:

-   -   i) at least one general formula (I) compound, preferably general        formula (II), (III) or (IV), or any combination thereof:    -   ii) pharmaceutically-acceptable vehicles; and    -   iii) additionally, a therapeutically-active substance

The term “pharmaceutically-acceptable vehicle” is understood as employedin the invention described herein as one or more excipients and/orcarrier substances or auxiliary substances which are pharmaceutically orpharmacological tolerable, such that they may be combined with othercomponents in the formulation or preparation and will have no adverseeffects on the organism treated.

The term “therapeutically-active substance” is understood as employed inthe invention described herein, as any substance synergisticallyinteracting with resveratrol, such as, but not limited to, otherpolyphenols such as quercetin.

The pharmaceutical compositions include, but are not limited to, thosewhich are suitable for oral or parenteral (including subcutaneous,intradermic, intramuscular and intravenous) administration, although thebest way of administering depends upon the patient's condition.

The formulations may be in single-dose form and be prepared inaccordance with methods known by any expert on the subject in the fieldof pharmacology. The quantities of active substances to be administeredmay vary in terms of the individual aspects of the therapy.

The following examples and figures provided in following serve toillustrate yet not limit the invention described herein.

EXAMPLES OF EMBODIMENT Example 1 Preparation of5-((E)-(4-Hydroxyphenylimine)methyl)benzene-1,3-diol, of the FollowingStructural Formula

Anhydrous magnesium sulfate was added to a suspension of drydichloromethane (5 ml) of 4-aminophenol (0.109 g, 1 mmol) and of 3,5dihydroxybenzaldehyde (0.138 g, 1 mmol). The resulting mixture wasagitated at ambient temperature for 3 hours. In following, the solventwas evaporated a low pressure and ethanol (5 mL) is added to theresulting residue. The mixture was heating to boiling and then filtered.The filtrate was evaporated under low pressure to yield a residue whichwas ground to a minimal quantity of cold ethanol, thus yielding5-((E)-(4-Hydroxyphenylimine)methyl)benzene-1,3-diol. Yield: 100%; p. f.(° C.) 162 (desc.).); IR (KBr) 3494, 3287, 1631, 1598, 1508, 1461, 1339,1273, 1155 cm⁻¹; ¹H NMR (300 MHz, DMSO-d₆) δ 9.44 (s, 1H), 9.42 (s, 2H),8.38 (s, 1H), 7.15 (d, J=7.8 Hz, 2H), ¹³C NMR (63 MHz, DMSO-d₆) δ 158.6,157.4, 156.2, 142.6, 138.4, 122.5, 115.7, 106.4, 105.3. Calc. Analysisfor C₁₃H₁₁NO₃: C, 68.11; H, 4.84; N, 6.11. Found: C, 68.76; H, 4.92; N,6.21%.

Example 2 Preparation of3,5-bis(3,5-dimethoxyphenyl)-1H-pyrrole-2-methyl carboxylate, of theFollowing Structural Formula

and of 3,5-bis(3,5-dimethoxyphenyl)-4-nitro-1H-pyrrole-2-methylcarboxylate, of the following structural formula:

In a spherical flask, (3,5-dimethoxybenzyllidenamine)methyl acetate (2.0g, 8.4 mmoles) was dissolved in 84 ml of CH₃CN, then adding 1.2 ml (1.25mmoles) de triethyllamine, 1,3-dimethoxy-5-(2-nitrovinyl)benzene (1.8 g,8.4 mmoles) and 0.21 g (1.25 mmoles) of AgOAc. The progress of thereaction was monitored by means of thin-layer chromatography. Followingcompletion of the reaction (approx. 5 hours), the mixture was filteredthrough a celite bed and the filtrate washed with an aqueous NH₄Clsolution (2×84 ml) and H₂O (2×84 ml), was then dried on anhydrous Na₂SO₄and evaporated at low pressure. The crude portion was purified by meansof pressurized chromatography column (AcOEt/Hx). 1.8 g (4 mmoles) of theoil obtained was dissolved in 40 ml of 2-methoxyethylether under argonatmosphere. In following, 3.5 g (40 mmoles) MnO₂ were added and agitatedto reflux for 48 hours. The reaction mixture was filtered through celiteand the filtrate evaporated at low pressure. The products were separatedby means of flash column chromatography, obtaining3,5-bis(3,5-dimethoxyphenyl)-1H-pyrrole-2-methyl carboxylate and3,5-bis(3,5-dimethoxyphenyl)-4-nitro-1H-pyrrole-2-methyl carboxylate.

3,5-bis(3,5-dimethoxyphenyl)-4-nitro-1H-pyrrole-2-methyl carboxylate:Yield. 25%; p.f. 139-141° C.; IR 3447, 3246, 1698, 1592, 1507, 1211,1161 cm⁻¹; ¹H-RMN (δ ppm, CDCl₃) 9.42 (s, 1H), 6.70 (d, 2H, J=2.3 Hz),6.56 (t, 1H, J=82.2 Hz), 6.50 (s, 3H), 3.82 (s, 6H), 3.69 (s, 3H);¹³C-RMN (δ ppm, CDCl₃) 161.3, 161.1, 160.5, 134.1, 133.9, 133.1, 130.6,124.4, 118.4, 108.3, 107.2, 102.6, 100.7, 55.9, 55.7, 52.5. Calc.Analysis for C₂₂H₂₂N₂O₈: C, 59.73; H, 5.02; N, 6.33. Found: C, 59.41; H,4.73; N, 6.36%.

3,5-bis(3,5-dimethoxyphenyl)-1H-pyrrole-2-methyl carboxylate: Yield: 31%p.f. 113-115° C.; IR 3276, 1668, 1608, 1156 cm⁻¹; ¹H-RMN (δ ppm, CDCl₃)9.28 (s, 1H), 6.77 (d, 2H, J=2.3 Hz), 6.71 (d, 2H, J=2.2 Hz), 6.60 (d,2H, J=3.1 Hz), 6.60 (d, 2H, J=3.1 Hz), 6.46 (t, 1H, J=2.3 Hz), 6.44 (t,1H, J=2.2 Hz), 3.84 (s, 6H), 3.82 (s, 3H); ¹³C-RMN (δ ppm, CDCl₃) 161.9,161.7, 160.5, 137.1, 135.7, 133.5, 133.1, 118.6, 110.6, 107.9, 103.4,100.4, 100.0, 55.9, 55.7, 51.9. Calc. Analysis for C₂₂H₂₃N₂O₆: C. 66.49;H. 5.84; N. 3.52. Found: C. 66.13; H. 5.47; N, 3.66%.

Example 3 Preparation of5-(3,5-dimethoxyphenyl)-3-(4-methoxyphenyl)-1H-pyrrole-2-methylcarboxylate, of the Following Structural Formula

and of5-(3,5-dimethoxyphenyl)-3-(4-methoxyphenyl)-4-nitro-1H-pyrrole-2-methylcarboxylate, of the following structural formula:

Both compounds were prepared and separated by means of a proceduresimilar to that described in Example 2 hereinabove.

5-(3,5-dimethoxyphenyl)-3-(4-methoxyphenyl)-4-nitro-1H-pyrrole-2-methylcarboxylate: Yield: 20%; p.f. 194-145° C.; IR 3266, 1683, 1597, 1507,1276, 1211, 1166 cm⁻¹; ¹H-RMN (δ ppm. CDCl₃) 9.42 (s. 1H). 7.32 (d. 2H.J=8.3 Hz). 6.96 (d. 2H. J=8.3 Hz). 6.71 (s. 2H). 6.58 (s. 1H). 3.87 (s.3H). 3.84 (s. 3H). 3.71 (s. 3H); ¹³C-RMN (δ ppm. CDCl₃) 161.4, 161.2,159.7, 134.4, 133.9, 131.4, 130.6, 127.6, 123.2, 118.3, 113.5, 107.2,103.5, 102.4, 55.9, 55.5, 52.4. Calc. Analysis for C₂₁H₂₀N₂O₇: C. 61.16;H. 4.90; N. 6.80. Found: C. 61.47; H. 4.92; N. 7.00%.

5-(3,5-dimethoxyphenyl)-3-(4-methoxyphenyl)-1H-pyrrole-2-methylcarboxylate: Yield: 35%; p.f. 123-125° C.; IR 3286, 1678, 1211 cm⁻¹;¹H-RMN (δ ppm, CDCl₃) 9.24 (s, 1H), 7.55 (d, 2H, J=8.6 Hz), 6.95 (d, 2H,J=8.6 Hz), 6.72 (s, 2H), 6.58 (d, 2H, J=2.9 Hz), 6.45 (s, 1H), 3.86 (s,9H), 3.82 (s, 3H); ¹³C-RMN (δ ppm, CDCl₃) 161.9, 161.7, 159.2, 135.7,133.7, 133.3, 130.8, 127.7, 118.2, 113.6, 110.4, 103.5, 100.3, 55.8,55.6, 51.7. Calc. Analysis for C₂₁H₂₁NO₅: C, 68.65; H, 5.77; N, 3.81.Found: C, 68.35; H, 6.00; N, 3.95%.

Example 4 Preparation of5-(3,5-dimethoxyphenyl)-3-(4-methoxyphenyl)-1H-pyrrole-2-methylcarboxylate, of the Following Structural Formula

and of5-(3,5-dimethoxyphenyl)-3-(4-methoxyphenyl)-4-nitro-1H-pyrrole-2-methylcarboxylate, of the following structural formula:

Both compounds were prepared and separated by means of a proceduresimilar to that described in Example 2.

3-(3,5-dimethoxyphenyl)-5-(4-methoxyphenyl)-4-nitro-1H-pyrrole-2-methylcarboxylate: Yield: 20%; p.f. 176-178° C.; IR 3286, 1683, 1618, 1507,1311 cm⁻¹; ¹H-RMN (δ ppm, CDCl₃) 9.20 (s, 1H), 7.56 (d, 2H, J=8.7 Hz),7.02 (d, 2H, J=8.7 Hz), 6.52 (s, 3H), 3.89 (s, 3H), 3.82 (s, 6H), 3.74(s, 3H); ¹³C-RMN (δ ppm, CDCl₃) 161.1, 160.2, 134.7, 133.2, 130.5,127.4, 120.9, 118.0, 114.4, 108.1, 100.4, 100.3, 55.6, 55.5, 52.2. Calc.Analysis for C₂₁H₂₀N₂O₇: C, 61.16; H, 4.90; N, 6.80. Found: C, 61.20; H,5.15; N, 6.87%.

3-(3,5-dimethoxyphenyl)-5-(4-methoxyphenyl)-1H-pyrrole-2-methylcarboxylate: Yield: 28%; p.f. 149-150° C.; IR 3326, 1673, 1151 cm⁻¹;¹H-RMN (δ ppm, CDCl₃) 9.18 (s, 1H), 7.51 (d, 2H, J=8.7 Hz), 6.96 (d, 2H,J=8.7 Hz), 6.77 (d, 2H, J=2.5 Hz), 6.53 (d, 2H, J=2.5 Hz), 6.45 (s, 1H),3.84 (s, 3H), 3.82 (s, 6H), 3.80 (s, 3H); ¹³C-RMN (δ ppm, CDCl₃) 161.8,160.3, 159.7, 135.1, 135.8, 133.5, 126.3, 123.9, 117.8, 114.6, 109.3,107.7, 99.8, 55.5, 51.5. Calc. Analysis for C₂₁H₂₁NO₅: C, 68.65; H,5.77; N, 3.81. Found: C, 68.52; H, 5.42; N, 4.08%.

Example 5 Preparation of3,5-bis(3,5-Dihydroxyphenyl)-1H-pyrrole-2-methyl carboxylate, of theFollowing Structural Formula

In a spherical flask cooled to 0° C. and under argon atmosphere, 0.2 g(0.5 mmoles) of 3,5-bis(3,5-Dimethoxyphenyl)-1H-pyrrole-2-methylcarboxylate (Prepared as stated in Example 2) were dissolved in 12 ml ofdry dichloromethane. In following, 6 ml of BBr₃ (1 M in dichloromethane)were added drop by drop and agitated to ambient temperature for 16hours. The reaction was halted by adding MeOH at 0° C. drop by drop. Theresulting solid was filtered and purified by means of pressurized columnchromatography. (Eluent: MeOH/CH₂Cl₂) obtaining the compound3,5-bis(3,5-hydroxyphenyl)-1H-pyrrole-2-methyl carboxylate.

3,5-bis(3,5-hydroxyphenyl)-1H-pyrrole-2-methyl carboxylate: Yield: 69%p.f. 233-235° C.; IR 3427, 1618 cm⁻¹; ¹H-RMN (δ ppm, DMSO-d₆) 11.2 (s,1H), 9.20 (s, 2H), 9.03 (s, 2H), 7.04 (s, 1H), 6.52 (s, 1H), 6.48 (s,2H), 6.40 (s, 2H), 6.09 (s, 1H), 6.02 (s, 1H); ¹³C-RMN (δ ppm, DMSO-d₆)161.0, 157.7, 157.2, 137.2, 136.3, 133.4, 127.1, 122.4, 117.1, 115.6,108.5, 107.7, 101.3, 51.0. Calc. Analysis for C₁₈H₁₅NO₆: C, 66.46; H,4.65; N, 4.31. Found: C, 66.13; H, 4.57; N, 4.36%.

Example 6 Preparation of5-(3,5-Dihydroxyphenyl)-3-(4-hydroxyphenyl)-1H-pyrrole-2-methylcarboxylate, of the Following Structural Formula

This compound was prepared and purified according to a procedure similarto that described in Example 5.

5-(3,5-Dihydroxyphenyl)-3-(4-hydroxyphenyl)-1H-pyrrole-2-methylcarboxylate: Yield: 63%; p.f. breaks down; IR 3537, 3397, 3316, 1673,1597, 1271, 1161 cm⁻¹; ¹H-RMN (δ ppm, DMSO-d₆) 11.6 (s, 1H), 9.41 (s,1H), 9.34 (s, 2H), 7.34 (d, 2H, J=8.5 Hz), 6.74 (d, 2H, J=8.5 Hz), 6.67(d, 2H, J=2.1 Hz), 6.46 (d, 2H, J=2.1 Hz), 6.19 (s, 1H), 3.68 (s, 3H);¹³C-RMN (δ ppm, DMSO-d₆) 161.0, 158.6, 156.5, 136.2, 132.9, 130.4,126.0, 117.4, 114.6, 109.4, 104.0, 102.2, 50.9. Calc. Analysis forC₁₈H₁₅NO₅: C, 66.46; H, 4.65; N, 4.31. Found: C, 66.60; H, 4.41; N,4.26%.

Example 7 Preparation of3-(3,5-Dihydroxyphenyl)-5-(4-hydroxyphenyl)-1H-pyrrole-2-methylcarboxylate, of the Following Structural Formula

This compound was prepared and purified according to a procedure similarto that described in Example 5.

3-(3,5-Dihydroxyphenyl)-5-(4-hydroxyphenyl)-1H-pyrrole-2-methylcarboxylate: Yield: 66%; p.f. 234-236° C.; IR 3326, 1678, 1608, 1281,1161 cm⁻¹; ¹H-RMN (δ ppm, DMSO-d₆) 11.6 (s, 1H), 9.64 (s, 1H), 9.16 (s,2H), 7.66 (d, 2H, J=8.6 Hz), 6.78 (d, 2H, J=8.6 Hz), 6.45 (d, 2H, J=2.3Hz), 6.37 (d, 2H, J=2.3 Hz), 6.19 (s, 1H), 3.69 (s, 3H); ¹³C-RMN (δ ppm,DMSO-d₆) 161.0, 157.7, 157.2, 137.2, 136.3, 133.4, 127.1, 122.4, 117.1,115.6, 108.5, 107.7, 101.3, 51.0. Calc. Analysis for C₁₈H₁₅NO₅: C,66.46; H, 4.65; N, 4.31. Found: C, 66.67; H, 4.61; N, 4.23%.

Example 8 Preparation of 3,5-bis(3,dimethoxyphenyl)-1H-pyrrole-2-carboxylic acid, of the FollowingStructural Formula

In a spherical flask, 0.45 g (1.1 mmoles) of3,5-bis(3,5-dimethoxyphenyl)-1H-pyrrole-2-methyl carboxylate (Preparedas stated in Example 2), 11 ml of 10% and 28 ml of ethanol were agitatedfor 3 hours to reflux. After cooling the reaction flask, the reactioncrude was neutralized with HCl 1M.

In following, the ethanol was evaporated at low pressure and theresulting aqueous phase was washed with AcOEt. Lastly, the organic phasewas dried on Na₂SO₄ and was evaporated at low pressure, obtaining thecompound 3,5-bis(3,5-dihydroxyphenyl)-1H-pyrrole-2-carboxylic acid.

3,5-bis(3,5-dimethoxyphenyl)-1H-pyrrole-2-carboxylic acid: Yield: 98%;p.f. 169-171° C.; IR 3316, 1653, 1608, 1196, 1156 cm⁻¹; ¹H-RMN (δ ppm,CDCl₃) 11.0 (s, 1H), 9.37 (s, 1H), 6.80 (d, 2H, J=2.0 Hz), 6.74 (d, 2H,J=2.0 Hz), 6.65 (d, 2H, J=2.8 Hz), 6.52-6.43 (m, 2H), 3.86 (s, 6H), 3.84(s, 6H); ¹³C-RMN (δ ppm, CDCl₃) 165.0, 161.6, 160.5, 136.7, 136.5,135.1, 132.7, 117.5, 110.9, 107.8, 103.5, 100.6, 100.2, 55.7, 55.6.Calc. Analysis for C₂₁H₂₁NO₆: C, 65.79; H, 5.52; N, 3.65. Found: C,65.71; H, 5.27; N, 3.55%.

Example 9 Preparation of3-(4-methoxyphenyl)-5-(dimethoxyphenyl)-1H-pyrrole-2-carboxylic acid, ofthe Following Structural Formula

This compound was prepared and purified according to a procedure similarto that described in Example 8.

3-(4-methoxyphenyl)-5-(dimethoxyphenyl)-1H-pyrrole-2-carboxylic acid:Yield: 85%; p.f. 152-153° C.; IR 3475, 3306, 1648, 1597, 1251, 1211,1156 cm⁻¹; ¹H NMR (δ ppm, CDCl₃) 11.3 (s, 1H), 9.30 (s, 1H), 7.56 (d,2H, J=8.5 Hz), 6.94 (d, 2H, J=8.5 Hz), 6.72 (d, 2H, J=1.7 Hz), 6.60 (d,1H, J=2.6 Hz), 6.46 (s, 1H), 3.85 (s, 9H); ¹³C NMR (δ ppm, CDCl₃) 161.2,158.7, 137.9, 133.2, 126.7, 125.7, 125.5, 115.3, 114.6, 103.7, 103.3,99.1, 55.5. Calc. Analysis for C₂₀H₁₉NO₅: C, 67.98; H, 5.42; N, 3.96.Found: C, 67.88; H, 5.51; N, 4.00;

Example 10 Preparation of 2,4-bis(3,5-dihydroxyphenyl)-1H-pyrrole, ofthe Following Structural Formula

0.1 g (0.26 mmoles) of the3,5-bis(3,5-dimethoxyphenyl)-1H-pyrrole-2-carboxylic acid (Prepared asstated in Example 8) were heated for 1 hour to 180-200° C. at a pressureof 1.2 mmHg. Upon completion of the reaction, the crude was cooled,obtaining 2,4-bis(3,5-dihydroxyphenyl)-1H-pyrrole, which was purified bymeans of column chromatography (Eluent: AcOEt/hexanes).

3,5-bis(3,5-Dimethoxyphenyl)-1H-pyrrole: Yield: 92%; p.f. 124-125° C.;IR 3427, 1597, 1212, 1161 cm⁻¹; ¹H-RMN (δ ppm, CDCl₃) 8.43 (s, 1H), 7.08(s, 1H), 6.75 (s, 1H), 6.70 (d, 2H, J=2.2 Hz), 6.63 (d, 2H, J=2.1 Hz),6.35 (t, 1H, J=2.1 Hz), 6.33 (t, 1H, J=2.2 Hz), 3.82 (s, 12H); ¹³C-RMN(δ ppm, CDCl₃) 161.5, 161.2, 137.7, 134.5, 133.1, 126.7, 116.0, 104.7,103.7, 102.5, 98.7, 98.1, 55.6, 55.5. Calc. Analysis for C₂₀H₂₁NO₄: C,70.78; H, 6.24; N, 4.13. Found: C, 70.65; H, 6.07; N, 4.20%.

Example 11 Preparation of4-amino-3,5-bis(3,5-dimethoxyphenyl)-1H-pyrrole-2-methyl carboxylate, ofthe Following Structural Formula

Seven milliliters (7 ml) of a SnCl₂ 2H₂O 1M in DMF were added to 0.2 g(0.45 mmol) of 3,5-bis(3,5-dimethoxyphenyl)-4-nitro-1H-pyrrole-2-methylcarboxylate (Prepared as stated in Example 2) and was agitated at 50° C.for 16 hours. Upon completion of the reaction, 4.5 ml of AcOEt and 10%aqueous Na₂CO₃ solution were added until no further precipitateappeared. The organic phase was separated and washed with saturatedNa₂CO₃ solution and water. After drying on anhydrous Na₂SO₄ andevaporating at low pressure, the crude portion was purified by means ofpressure column chromatography (Eluent: AcOEt/hexanes) obtaining4-amino-3,5-bis(3,5-dimethoxyphenyl)-1H-pyrrole-2-methyl carboxylate.

4-Amino-3,5-bis(3,5-dimethoxyphenyl)-1H-pyrrole-2-methyl carboxylate:Yield: 81%; p.f. 74-76° C.; IR 3417, 3346, 1678, 1597, 1211, 1151 cm⁻¹;¹H-RMN (δ ppm, CDCl₃) 8.74 (s, 1H), 6.74 (d, 2H, J=2.2 Hz), 6.60 (d, 2H,J=2.1 Hz), 6.46 (t, 1H, J=2.1 Hz), 6.40 (t, 1H, J=2.0 Hz), 3.84 (s, 6H),3.81 (S, 6H), 3.73 (s, 3H); ¹³C-RMN (δ ppm, CDCl₃) 161.6, 160.6, 135.0,133.6, 129.5, 121.6, 120.5, 116.7, 108.2, 103.7, 99.8, 99.2, 55.6, 55.5,51.5. Calc. Analysis for C₂₂H₂₄N₂O₆: C, 64.07; H, 5.87; N, 6.79. Found:C, 63.91; H, 5.74; N, 6.57%.

Example 12 Preparation of4,6-dimethoxy-2-(3,5-dimethoxyphenyl)-1H-indole, of the FollowingStructural Formula

0.32 g (2.1 mmoles) of 3,5-dimethoxyaniline, 0.26 g (1.0 mmol) of2-bromo-1-(3,5-dimethoxyphenyl)ethanone and 0.42 ml (3.3 mmol) ofN,N-dimethylaniline were placed in a vial. The vial was placed inside amonomode microwave reactor and was radiated at a power of 100 W, at atemperature of 150° C. for 10 minutes. After cooling the reaction vial,the mixture was dissolved in AcOEt, was washed with an aqueous HCl 2Nsolution, was dried on anhydrous Na₂SO₄ and was evaporated at lowpressure. The crude portion was purified by means of pressure columnchromatography (Eluent: AcOEt/hexanes) obtaining4,6-dimethoxy-2-(3,5-dimethoxyphenyl)-1H-indole.

4,6-dimethoxy-2-(3,5-dimethoxyphenyl)-1H-indole: Yield: 80%; p.f.126-128° C.; IR 3407, 1608, 1206, 1161 cm⁻¹; ¹H-RMN (δ ppm, CDCl₃) 8.18(s, 1H), 6.82 (s, 1H), 6.73 (d, 2H, J=1.8 Hz), 6.49 (s, 1H), 6.38 (s,1H), 6.22 (s, 1H), 3.92 (s, 3H), 3.83 (s, 9H); ¹³C-RMN (δ ppm, CDCl₃)161.4, 158.2, 153.9, 138.3, 135.3, 134.7, 114.6, 103.1, 99.4, 97.8,92.2, 87.1, 55.8, 55.6. Calc. Analysis for C₁₈H₁₉NO₄: C, 68.99; H, 6.11;N, 4.47. Found: C, 68.92; H, 5.84; N, 4.46%.

Example 13 Preparation of4,6-dimethoxy-2-(2,5-dimethoxyphenyl)-1H-indole, of the FollowingStructural Formula

The compound stated in the heading above was prepared and purifiedaccording to a procedure similar to that described in Example 11.4,6-dimethoxy-2-(2,5-dimethoxyphenyl)-1H-indole: Yield: 61%; p.f.148-150° C.; IR 3407, 1542, 1487, 1211, 1141 cm⁻¹; ¹H-RMN (δ ppm, CDCl₃)9.62 (s, 1H), 7.32 (d, 2H, J=3.0 Hz), 6.92 (d, 2H, J=8.9 Hz), 6.90 (d,1H, J=1.3 Hz), 6.76 (dd, 1H, J=8.9, J′=3.0 Hz), 6.52 (s, 1H), 6.21 (d,1H, J=1.3 Hz), 3.94 (s, 6H), 3.84 (s, 3H), 3.81 (s, 3H); ¹³C-RMN (δ ppm,CDCl₃) 157.7, 154.3, 153.5, 149.9, 137.4, 133.1, 121.7, 113.5, 113.4,112.3, 97.2, 91.7, 86.8, 56.6, 55.8, 55.7, 55.4. Calc. Analysis forC₁₈H₁₉NO₄: C, 68.99; H, 6.11; N, 4.47. Found: C, 68.72; H, 6.44; N,4.45%.

Example 14 Preparation of4,6-dimethoxy-2-(3,5-dimethoxyphenyl)-1H-indole-1-tert-butylcarboxylate, of the Following Structural Formula

0.20 g (0.92 mmol) of Boc₂O and 0.009 g (0.071 mmol) of4-dimethylaminopyridine were added to a solution of 0.21 g (0.68 mmol)of 4,6-dimethoxy-2-(3,5-dimethoxyphenyl-1H-indole (Prepared according tothe procedure described in Example 11) in 8.2 ml of acetonitryl and wasagitated at ambient temperature for 3 hours. In following, the solventwas evaporated at reduced pressure and the resulting product waspurified by means of pressure column chromatography (Eluent:AcOEt/hexanes), obtaining the compound4,6-dimethoxy-2-(3,5-dimethoxyphenyl)-1H-indole-1-tert-butylcarboxylate.

4,6-Dimethoxy-2-(3,5-dimethoxyphenyl)-1H-indole-1-tert-butylcarboxylate: Yield: 60%; p.f. 92-94° C.; IR 1733, 1613, 1587, 1311, 1151cm⁻¹; ¹H-RMN (δ ppm, CDCl₃) 7.38 (d, 1H, J=1.6 Hz), 6.59 (s, 1H), 6.54(d, 1H, J=2.3 Hz), 6.44 (t, 1H, J=2.3), 6.35 (d, 1H, J=1.6 Hz), 3.89 (s,3H), 3.88 (s, 3H), 3.79 (s, 6H), 1.33 (s, 9H); ¹³C-RMN (δ ppm, CDCl₃)160.4, 159.3, 153.3, 150.6, 139.3, 137.6, 137.2, 113.8, 107.1, 106.8,99.9, 94.8, 91.6, 83.5, 56.0, 55.6, 27.8. Calc. Analysis for C₂₃H₂₇NO₆:C, 66.81; H, 6.58; N, 3.39. Found: C, 66.67; H, 6.44; N, 3.44%.

Example 15 Preparation of4,6-dimethoxy-2-(2,5-dimethoxyphenyl)-1H-indole-1-tert-butylcarboxylate, of the Following Structural Formula

The compound stated in the heading above was prepared and purified bymeans of a procedure similar to that described in Example 13.

4,6-Dimethoxy-2-(2,5-dimethoxyphenyl)-1H-indole-1-tert-butylcarboxylate: Yield: 86%; p.f. 56-58° C.; IR 1783, 1497, 1311, 1221, 1151cm⁻¹; ¹H-RMN (δ ppm, CDCl₃) 7.39 (d, 1H, J=1.5 Hz), 6.94 (d, 1H, J=3.0Hz), 6.84 (dd, 1H, J=8.8 Hz, J′=3.0 Hz), 6.77 (d, 1H, J=8.8), 6.54 (s,1H), 6.33 (d, 1H, J=1.5 Hz), 3.88 (s, 6H), 3.78 (s, 3H), 3.67 (s, 3H),1.30 (s, 9H); ¹³C-RMN (δ ppm, CDCl₃) 159.0, 153.6, 153.2, 151.7, 150.5,138.7, 134.2, 125.9, 116.2, 113.6, 111.1, 106.5, 94.4, 91.5, 82.8, 56.0,55.9, 55.6, 27.7. Calc. Analysis for C₂₃H₂₇NO₆: C, 66.81; H, 6.58; N,3.39. Found: C, 66.81; H, 6.24; N, 3.51%.

Example 16 Preparation of 2-(2,5-dihydroxyphenyl)-1H-indole-4,6-diol, ofthe Following Structural Formula

In a spherical flask cooled to 0° C. and under argon atmosphere, 0.2 g(0.5 mmol) of4,6-dimethoxy-2-(2,5-dimethoxyphenyl)-1H-indole-1-tert-butyl carboxylate(Prepared as described in Example 14) were dissolved in 12 ml of drydichloromethane. In following, 6 ml of BBr₃ (1 M in dichloromethane)were added drop by drop and agitated to ambient temperature for 16hours. The reaction was halted by adding MeOH drop by drop at 0° C. Theresulting solid was filtered and was purified by means of pressurecolumn chromatography (Eluent: MeOH/CH₂Cl₂) obtaining the compound2-(2,5-dihydroxyphenyl)-1H-indole-4,6-diol.

2-(2,5-Dihydroxyphenyl)-1H-indole-4,6-diol: Yield: 52%; p.f. breaksdown; IR 3363, 1612, 1456, 1207 cm⁻¹; ¹H-RMN (δ ppm, DMSO-d₆) 10.46 (s,1H), 9.14 (s, 1H), 9.09 (s, 1H), 8.65 (s, 1H), 8.60 (s, 1H), 7.01 (d,1H, J=2.8 Hz), 6.80 (d, 1H, J=1.3 Hz), 6.73 (d, 1H, J=8.6 Hz), 6.60 (dd,1H, J=8.6 Hz, J′=2.8 Hz), 6.29 (s, 1H), 5.92 (d, 1H, J=1.3 Hz); ¹³C-RMN(δ ppm, DMSO-d₆) 153.9, 150.3, 149.8, 146.3, 138.4, 131.3, 119.8, 116.8,113.8, 112.5, 112.2, 98.4, 94.5, 88.2. Calc. Analysis for C₁₄H₁₁NO₄: C,65.37; H, 4.31; N, 5.44. Found: C, 65.52; H, 4.36; N, 5.11%.

Example 17 Tests Conducted to Evaluate the In Vitro and In VivoBiological Activity of the General Formula (I) Compounds

The potential antimetastatic effect of general formula (II) compoundswas studied by using an in vivo experimental model of inflammation andoxidative stress-dependent hepatic colonization ofintrasplenically-injected B16 melanoma (MB16) cells, in which themetastasis density and volume, the type of metastasis based on theangiogenic (sinusoidal or portal) pattern thereof, the length of theangiogenic vessels within tumor nodules, the percentage of metastaticfocal points with cavitation and the percentage of in situ proliferatingmetastatic cells were histologically determined. In following, for thepurpose of studying in what stage of the development of the metastasissaid compounds were acting, the in vivo effect thereof on the hepaticretention of the tumor cells and in vitro, on one hand, on theproduction of hydrogen peroxide, adhesion and proliferation of murineB16 melanoma and human A375 melanoma cells and, on the other hand, onthe migration and proliferation of primary cultures of endothelial cellsand hepatic estellate cells was determined.

Following an optimization of the chemical structure, the general formula(III) compounds and the general formula (IV compounds, the chemical andconfigurational stability of which afford the possibility of preservingthe trans arrangement between the aromatic rings (A and B), see generalformula (I), avoiding the possibility of isomerization and the resultingloss activity. The following in vitro tests were conducted with thesecompounds:

Tests of B16 (MB16) melanoma cell interaction with primary cultures ofhepatic sinusoidal endothelium (HSE cells treated with conditioned MB16(MC-MB16) media:

-   -   ELISA determination of the TNF-alpha concentration in the        supernatants of HSE cell cultures treated with MC-MB16.    -   Determination of the production of hydrogen peroxide from HSE        cells and MB16 cells.    -   Cell adhesion tests of MB16 cells incubated with non-toxic        concentrations of hydrogen peroxide to the immobilized        recombinant VCAM-1 substrate.    -   Proliferation tests of the MB16 cells treated with recombinant        murine IL-18.    -   Enzymatic immunoassay (EIA) of PGE2 in the supernatant of the        untreated and VEGF-treated HSE cells.

Lastly, an evaluation was made on the effect of those compounds showinggreater inhibitory activity in the in vitro tests on the metastaticcapacity of the MB16 cells by means of in vivo tests on the developmentof hepatic metastasis.

Example 17.a) Tests Conducted to Evaluate the In Vitro BiologicalActivity

Culture of B16 (MB16) and A375 (MA375) melanoma cells. The MB16 (B16F10subline) murine tumor cells were cultured at 37° C., with 5% CO₂atmosphere in Dulbecco's modified Eagle's medium (DMEM)-HCO₃—,penicillin (100 U/ml), streptomycin (100 μg/ml), supplemented with 5%fetal bovine serum and adjusted to pH 7.4. The cells were maintained andsubcultured according to the method described by Vidal-Vanaclocha et al.(1994) [Vidal-Vanaclocha, F., Amézaga, C., Asumendi, A., Kaplanski, G. &Dinarello, C. A. Interleukin-1 receptor blockade reduces the number andsize of murine B16 melanoma hepatic metastases. Cancer Research, 1994,54, 2667-2672]. The conditioned media were obtained from subconfluyentcultures maintained in absence of FBS for 24 hours.

Isolation of Primary murine cultures of hepatic sinusoidal endothelial(HSE) cells. The hepatic sinusoidal cells were isolated from C57BL/6Jmice (males, aged 6-8 weeks) supplied by IFFA Credo (L'Arbreole,France), followed by a purification and identification of the HSE cellsfollowing the protocol described by Vidal-Vanaclocha et al. (1993)[Vidal-Vanaclocha, F., Rocha, M., Asumendi, A. & Barbera-Guillem, E.Isolation and enrichment of two sublobular compartment-specificendothelial cell subpopulations from liver sinusoids. Hepatology, 1993,18, 328-339]. Stable primary cultures of HSE cells were obtained afterseeding the cells on a type I collagen substrate (Sigma Chemicals Co, StLouis, Mo.), and the cultures were maintained in DMEM-HCO³⁻, penicillin(100 U/ml), streptomycin (100 μg/ml), supplemented with 10% fetal bovineserum at 37° C. with 5% CO2 in atmosphere at 98% humidity.

Isolation of primary human cultures of hepatic sinusoidal endothelial(HSE) hepatic stellate cells (HSCs). The primary cultures of human HSEand HSC cells were obtained by non-tumoral hepatic tissue serialprofusion with collagenase and pronase solutions, followed by Nycodenzgradient and seeded on 24-well plates (0.5×104 cells/cm²) in DMEMsupplemented with 10% FBS.

MB16 cell adhesion to the monolayer of HSE cells in culture. Theendothelial cells were isolated 24 hours prior to the adhesion test andwere seeded on 24-well plate, maintaining them a minimum of 4 hours inserum-free medium before incubating them with DMEM in the presence ornot of MC-MB16 for 8 hours. The synthesized compounds (JE2:2 see FIG. 6;JEM2:2-01 and JEM2:1-02 see FIG. 8A; YEF02, YEF03, YEF07, YEF05B, YEF07Band YEF05H see FIG. 10) were added to a 2.5 μM concentration prior tothe MC-MB16. On the other hand, the MB16 cells were marked with 40 μg/mlfrom the fluorescent probe BCECF-AM (carboxyfluorescein,2′,7′-bis-(2-carboxy-ethyl)-5-(6)carboxyfluorescein aminoxymethyl ester)supplied by Molecular Probes Inc. (Oregon, USA). Afterward, a washingprocess was performed with DMEM-HCO₃— to remove the excessfluorochromium, the number of viable cells being calculated by means ofthe trypan blue exclusion test and resuspended to a concentration of2×10⁵ cells/ml. Lastly, 1 ml of the MB16 cell suspension was added toeach well on the primary HSE cell culture plate. The co-culture plateswere incubator-incubated at 37° C. for 8 minutes. The percentage ofcellular adhesion was calculated by means of the a fluorescencemeasurement system described by Vidal-Vanaclocha et al. (1994)[Vidal-Vanaclocha, F., Amézaga, C., Asumendi, A., Kaplanski, G. &Dinarello, C. A. Interleukin-1 receptor blockade reduces the number andsize of murine B16 melanoma hepatic metastases. Cancer Research, 1994,54, 2667-2672]. In the experiments with IL-18, the MB16 cells wereincubated with 10 ng/ml of IL-18 for 4 hours prior to being marked withBCECF-AM and were adhered to the HSE cell plate. The results in thefigures are given as values related to the adhesion percentages of theuntreated HSE cells.

MB16 and MA375 cell adhesion to immobilized VCAM-1 substrates. The MB16and MA375 cell adhesion tests were conducted on immobilized VCAM-1substrates (2 μg/ml of recombinant human VCAM-1, R&D Systems,Minneapolis, Minn.) on 96-well plates. To avoid non-specific attachmentto the plastic, 0.5% BSA dissolved in PBS was added to the wells for 2hours at ambient temperature prior to conducting the adhesion test. Themelanoma cells were preincubated with 2.5 μM of the compounds to betested (azaresveratrol, see Table 2; JE1:2, JE2:1 and JE2:2 see FIG. 2;JEM2:2-01 and JEM2:1-02 see FIG. 7; YEF02, YEF03, YEF07, YEF05B, YEF07Band YEF05H see FIG. 9) for 30 minutes, to which 10 μM of hydrogenperoxide or 10 ng/ml of IL-18 were added, respectively for a further 24hours. Afterward, the cells were washed and marked with the BCECF-AMfluorescent probe (carboxyfluorescein,2′,7′-bis-(2-carboxy-ethyl)-5-(6)carboxyfluorescein aminoxymethylester). Thirty minutes later, a washing process was performed to removethe excess fluorochromium, the number of viable cells being calculatedby means of the trypan blue exclusion test and (5×10⁴ cells/well) beingadded to the 96-well plate. The co-culture plates wereincubator-incubated at 37° C. for 1 hours. The percentage of cellularadhesion was calculated by measuring the fluorescence emitted by theadhered cells (obtained after washing the plate) with regard to thefluorescence emitted by the total number of cells added. The results inthe figures are given in values related to the adhesion percentages ofthe untreated melanoma cells.

ELISA determination of the TNF-alpha concentration in the supernatantfrom MC-MB16-treated HSE cells. Primary cultured HSE cells wereincubated in the presence or absence of 2.5 μM of the synthesizedcompounds (JEM2:2-01 and JEM2:1-02 see FIG. 8B) for 30 minutes, afterwhich MB16-conditioned media were added. Eight (8) hours later, thesupernatant from cultured endothelial cells were collected, filteredthrough 0.22 μm membranes and the concentration of TNF-alpha was thenassayed using the ELISA test (R&D Systems).

Determination of hydrogen peroxide production from HSE and MB16 cells.Primary cultures of HSE cells were treated with 10 μg/ml of the DCFH-DAfluorescent probe for 30 minutes at 37° C. and were washed to eliminatethe excess fluorochromium. Afterward, fresh culture medium was addedthereto in the presence or absence of 2.5 μM of the compounds JE1:2,JE2:1 or JE2:2 and the production of H202 over the course of time wasdetermined (see FIG. 5). The hydrogen peroxide produced by the cellsoxidizes the probe, converting it into a fluorescent molecule. Thus, thecell fluorescence produced by the accumulation of DCF made it possibleto detect the intracellular production of hydrogen peroxide. In eachcell well, it was possible to determine the relative fluorescence value(arbitrary units of fluorescence brightness) in relation to the quantityof hydrogen peroxide produced at different incubation times. In theexperiments with MB16 cells, once the cells had been marked with theDCFH-DA probe, they were incubated for 300 minutes with 2.5 μM of theazaresveratrol or JE2:2 compounds, 10 ng/ml of IL-18 (FIG. 4) thenhaving been added.

In vitro Melanoma cell proliferation test. The MB16 or MA375 cells areseeded (2500 cells/well/200 μl) in DMEM medium (Dulbecco's modifiedEagle's medium) with 10% fetal bovine serum (FBS). Once adhered, theywere washed to remove the FBS and fresh medium was added in the presenceor absence of 2.5 μM of the compounds to be tested. In the experimentswith MA375, the compound azaresveratrol (see Table 4) was used, and inthe experiments with MB16 cells, the compounds azaresveratrol, JE1:2,JE2:1 or JE2:2 (see FIG. 3) were used. Thirty (30) minutes later, 10ng/ml of IL-18 or HSC-conditioned media (murines in the case of B16 andhuman ones in the case of A375) were added to some wells, and the plateswere incubated for 48 hours at 37° C. with 5% CO₂ Upon completion of theincubation period, the cells were fixed with 64% for 1 hours, werewashed and were allowed to dry in the incubator at 50° C. for 30minutes. In following, 100 μl/well of sulforhodamine 101 (0.4% p/v) wasadded and they were incubated at room temperature in the dark for 30minutes. After washing, 200 μl/well of Tris base 10 mM pH 10.5 was addedand the fluorescence was measured by means of a 530 nm, 620 nm emissionexcitation filter plate fluorescence reader. The number of cells wascalculated by extrapolating said fluorescence data on a standardstraight line previously obtained based on the fluorescence emitted inan increasing number of cells.

TABLE 4 IN VITRO TESTS CONTROL TREATMENT +RESVERATROL +AZARESVERATROLProliferation of MA375 cells in response to human 9000 ± 60 17500 ± 50  9050 ± 100 9250 ± 65 HSC soluble factors (No. cells per well)Proliferation of MB16 cells in response to murine 16300 ± 300 21660 ±500 16800 ± 250 16750 ± 135 HSC soluble factors (No. cells per well)

HSC proliferation test. Primary cultured human HSC cultures were seeded(2500 cells/well/200 μl) in DMEM with 10% FBS. Once adhered, they werewashed to remove the FBS and were incubated with fresh medium in thepresence or absence of 12.5 μM of the azaresveratrol compound (see Table3). Thirty (30) minutes later, conditioned media obtained from MA375cells were added to some wells, and the plates were incubated for 48hours in humid incubator at 37° C. with 5% CO₂. Upon completion of theincubation time, the cells were fixed and were processed for countingjust as previously described hereinabove.

PGE2 determination in HSE cell supernatants (see FIG. 11). Primarycultured HSE cells were incubated in the presence or absence ofincreasing concentrations (1, 2.5 and 10 μM) of the compounds JE2:2,YEF07 and YEF05B for 30 minutes. In following, 10 ng/ml of recombinantmurine VEGF or the same volume of saline solution was added, 4 hoursafter which supernatants were collected and the PGE2 concentrationdetermined by enzymatic immunoassay (EIA) supplied by AmershamBiosciences (Uppsala, Sweden).

Human HSC and HSE cell migration tests (see Table 3). Primary culturedhepatic endothelial cells (25×10⁵) or hepatic stellate cells (2×10⁴)were seeded in the upper compartment of modified Boyden chambersequipped with a polycarbonate filter of an 8 μm pore size. Some cellswere incubated n the presence or absence of 12.5 μM of azaresveratrol,30 minutes prior to adding conditioned tumor medium. Twenty-four (24)hours later (in the case of the endothelial cells) or 4 hours later (inthe case of hepatic star cells) the number of cells having passedthrough the membrane was determined. The cells were fixed, were stainedwith hematoxylin eosine and were counted under the microscope (20×) in 5fields per well.

TABLE 3 IN VITRO TESTS CONTROL TREATMENT +RESVERATROL +AZARESVERATROLMigration of human HSC's treated with MC- 10.0 ± 4  91.5 ± 19  6.5 ± 1.4  6.0 ± 1.3 MA375 (No. cells migrated per field) Proliferation of humanHSC's treated with MC-  2550 ± 22 4400 ± 30 2250 ± 100 2600 ± 60  MA375(No. cells per well) Migration of HSE's treated with MC- 10.0 ± 4  35.0± 8.0 13.5 ± 5.0 15.5 ± 30 MA375 (No. cells per field)

Example 17.b) Tests Conducted to Evaluate the In Vivo BiologicalActivity

MB16 Hepatic Metastasis Test using C57 BL/6J mice (males 6-8 weeks ofage) supplied by IFFA Credo (L'Arbreole, France). The care, maintenanceand experimental conditions were carried out in accordance with thatwhich is set forth under EEC Council Directive 86/609 (OJ L 358. 1, Dec.12, 1987) and the NIH guide for the care and use of laboratory animals(NIH publication 85-23, 1985). In the experiments with the resveratroland azaresveratrol compounds (Table 1), the animals (10 per group)received 1 mg/Kg/day of the compounds nasogastrically every day up tothe time of sacrifice. In the experiments with the JE2:2 compound (FIG.12), one group of mice (n=10) was given an intraperitoneal injection of0.5 mg/kg of the JE2:2 compound dissolved in 0.1 ml of PBS. The controlgroup (n=10), was given an intraperitoneal injection of PBS. One hourlater, animals were anesthetized and were intrasplenically injected withMB16 cells (3×10⁵ cells/mouse). The same treatments, at the same doses,were repeated on days 2, 3, 4, 8, 9, 10 and 11 following tumorinjection. Twelve (12) days later, the livers were removed and processedfor the histological study. Firstly, they were fixed in a solution ofzinc (0.5 g calcium acetate, 5 g zinc acetate, 5 g zinc chloride and1000 ml tris buffer, pH 7.4) for 24 hours. Once fixed, they weredehydrated in alcohols of increasing concentrations, and paraffin blockwere included. Afterward, a minimum of nine (9) cuts 10 μm in thicknesswere made per liver, leaving a space of 300 μm between every 3 cuts anda 1 mm space between successive groups of 3 cuts. Once the sections hadbeen obtained, they were processed and stained with hematoxylin-eosine.On the one hand, the number, mean diameter, area and positioncoordinates of each metastasis were quantified by means of an integratedimage analysis system (Olympus Microimage 4.0 capture kit) connected toan Olympus BX51TF microscope. With obtained data, metastatic parameters(that is to say, the number of focal points per 100 mm³ of hepatictissue) and metastatic volume (that is to say, the volume of liveroccupied by metastatic tissue) were calculated as described byVidal-Vanaclocha et al. [cf. Vidal-Vanaclocha, F., Amézaga, C.,Asumendi, A., Kaplanski, G. & Dinarello, C. A. “Interleukin-1 receptorblockade reduces the number and size of murine B16 melanoma hepaticmetastases”, Cancer Research, 1994, vol. 54, pp. 2667-2672]. On theother hand, more cuts were made in the same blocks, and a double markingwith CD31 and desmine was performed to calculate the number and lengthof the angiogenic vessels within the metastases and also to studymetastases according to their angiogenic pattern in portal-typeexpansive metastases (the angiogenic vessels surround the metastases) orsinusoidal type invasive metastases (containing an internal network ofangiogenic capillaries). Another parameter that was quantified is thecavitated metastasis percentage. Lastly, immunohistochemical detectionof Ki67 antigen was performed to assess the number of tumor cells whichproliferated per unit of tumor surface.

Test of hepatic retention of luciferase-transfected MB16 cells(MB16-Luc) (see Table 1). B16M Cells were stably transfected bylipofection as described previously (Rubio N, Martinez-Villacampa M,Blanco J. Traffic to lymph nodes of PC-3 prostate tumor cells in nudemice visualized using the luciferase gene as a tumor cell marker. LabInvest 1998; 78:1315-1325), using plasmidpRc/cytomegalovirus-luciferase, a construct containing the Photinuspyralis luciferase gene coding sequence under transcripcional control ofthe cytomegalovirus promoter and the neomycin resistance gene to theG418 antibiotic (Sigma Chemicals Co.). A total of 300,000 viableB16M-Luc cells were intrasplenically injected into C57BL/6J mice (10mice per group) which had previously been nasogastrically administered 2doses of 1 mg/kg/day of compound to be tested. One group of micereceived resveratrol, the other group of mice received azaresveratrol.The group of control mice were administered the same solution of thecompounds. All mice were killed by cervical dislocation 18 h later andthe livers were processed as described previously (Rubio et al, 1998) tomeasure luciferase activity by chemiluminiscence using the standardluciferase assay kit (Promega Co., Madison, Wis.) as reported (Rubio etal, 1998). Production of light was measured using a luminometer designedto read individual samples tubes (bioorbit, LKB Wallac; Turku, Finland)after the addition of 100 μl of luciferase assay reagent to 20 μl ofeach liver homogenate. Light detector measurements were expressed inrelative light units, which were proportional to photon numbers.Linearity and sensitivity of light detection in liver homogenates andinfluence of hepatic microenvironment on luciferase activity of B16M-Lucwere also evaluated as described previously (Rubio et al, 1998).

TABLE 1 IN VIVO TESTS CONTROL +RESVERATROL +AZARESVERATROL Hepaticretention of MB16 Luc cells  1.80 ± 0.23 1.05 ± 0.3   0.98 ± 0.24 (No.cells × 10⁶) Hepatic focal point density (No. focal 60.0 ± 11  30.0 ±11  28.0 ± 10 points/100 mm³) No. portal metases/No. sinusoidal 30/30(1) 22.5/7.5 (3) 22.5/5.5 (4.09) metastasis Volume of hepatic metastasis(mm³) 60.0 ± 7.5 18.7 ± 7.5  24.0 ± 9.0 Length of angiogenic vessels(μm) 175.0 ± 10   88.0 ± 23  88.0 ± 21 Percentage of focal pointsw/cavitation 13.5 ± 5.0 41.5 ± 11  40.0 ± 10 (% of total no. ofmetastatic focal points) Metastatic cell proliferation index 71.0 ± 5.040.0 ± 5.0 35.0 ± 10 (% cells positive to Ki67 antigen)

Example 17. c) General Formula (II) Compound Biological Activity ResultsIn Vivo Test Results:

Firstly, a comparative study was made between the effect of theresveratrol and the general formula compound (azaresveratrol) on theintrasinusoidal retention of the tumor cells throughout the first 18hours and their capacity to develop metastasis on day 12 following theirinoculation (see Table 1).

Both treatments significantly (P<0.05) reduced the intrahepaticretention of tumor cells 18 hours following their injection. Theseretention experiments were conducted with MB16 cells transfected withthe lucerifase gene (MB16-Luc precisely as previously described in themethods employed. In addition thereto, the daily administration of 1mg/Kg/day of resveratrol and azaresveratrol nasogastrically reduced themean density of hepatic metastases by 50% and 46%, respectively, whichdenotes a statistically significant (P<0.01) antimetastatic effect incomparison to the control animals. The immunohistochemical staining forCD31 and desmine revealed that the treatments affected to a greaterdegree the metastases with a sinusoidal-type angiogenic pattern, alsoknown as invasive metastases, whilst they had almost no effect on thenumber of metastases having a portal-type angiogenic pattern, orexpansive metastases. On the other hand, the percentage of hepaticvolume occupied by metastatic tissue decreased by 40% in the treatedanimals. Said inhibition in the volume had a reduction in the length ofangiogenic vessels, a larger percentage of metastatic foci with internalcavitation and a lower tumor cell proliferation index as determined byimmunohistochemistry for Ki67 antigen (see Table 1). These resultsprove, on one hand, that the general formula compound (azaresveratrol)has an in vivo antimetastatic efficiency similar to natural resveratroland, on the other hand, that the effect is two-way, affecting both thestroma as well as the tumor cells directly.

In Vitro Test Results:

In following, a series of in vitro tests are conducted for identifyingthe mechanisms of action of the azaresveratrol on the different stagesof the development of the metastases. The results are compared inrelation to those obtained with resveratrol.

Table 2 shows that the preincubation of the MB16 cells with 2.5 μM ofresveratrol and azaresveratrol for 30 minutes prior to adding IL-18significantly (P<0.01) inhibited the increase of both the percentage ofadhesion of tumor cells to the HSE and the production of tumoral H202 inresponse to IL-18. Not statistically significant differences were foundbetween the two compounds. Likewise, this inhibitory effect was alsoobserved in adhesion tests of A375 human melanoma cells incubated with10 μM H2O2 for 2 hours or with 1 ng/ml IL-18 for 6 hours to immobilizedVCAM-1 substrates. Therefore, the azaresveratrol inhibited earlyprocesses to the metastatic implantation such as tumoral adhesion to thevascular endothelium.

To determine whether the compounds affect mechanisms associated withmetastatic development and growth, migration tests are conducted onprimary human cultures of hepatic stellate cells (HSC) and hepaticsinusoidal endothelial cells (HSE) in response to soluble factors fromMA375 cells (see Table 3). When the HSE and HSC cells were administered12.5 μM both of resveratrol and azaresveratrol 30 min prior to theMC-A375, the increase in migration induced by the tumor was abolished.On the other hand, the same treatment abrogated the increase in HSCproliferation in response to the inhibition of the migration of HSEcells and HSCs as well as of the proliferation of the latter of the twoexplains the antiangiogenic effect of the compounds observed in the invivo experiments.

An evaluation was also carried aout to determine the direct effect ofthe resveratrol and azaresveratrol on the proliferation of B16 and A375cells in response to soluble factors secreted by HSCs. As shown in Table4, the treatment with MC-CEH for 48 hours significantly (P<0.01)increased the proliferation of melanoma cells. This increase in theproliferation was prevented by incubating the cells with 2.5 μM of bothresveratrol and azaresveratrol for 30 minutes before adding theMC-CEH's.

TABLE 2 IN VITRO TESTS CONTROL TREATMENT +RESVERATROL +AZARESVERATROLAdhesion of MB16 cells treated with IL-18 to 24.4 ± 4   54 ± 6.0  30 ±4.0  31 ± 4.0 HSE cells (% adhered cells) H₂O₂ production by MB16 cellstreated with IL-18 16.0 ± 3   34 ± 3.0  12 ± 1.0  10 ± 2.0 Adhesion ofMA375 cells with H₂O₂ to immobilized 21.0 ± 4 41.3 ± 4.2 6.5 ± 0.5 4.5 ±0.3 VCAM-1 (% cells adhered) Adhesion of MA375 cells treated with IL-18to 21.0 ± 4 31.5 ± 3.2 5.5 ± 0.4 5.0 ± 0.3 immobilized VCAM-1 (% cellsadhered)

Example 17. d) Biological Activity Results for General Formula (III) and(IV) Compounds In Vitro Test Results:

Firstly, the effect of the formula (III) compounds on the prometastaticbehavior of the tumor cells was studied. In FIG. 2, it was observed thatthe preincubation of the MB16 cells with the JE1:2, JE2:1 and JE2:2compounds significantly (P<0.01) inhibits the increase in the percentageof H2O2-treated melanoma cell adhesion to the immobilized VCAM-1. TheJE2:2 compound is the one showing the greatest inhibitory effect.

In the proliferation tests (see FIG. 3), the treatment with 10 ng/ml ofthe proinflammatory cytokine IL-18, produced a statistically significant(P<0,01) increase in the number of MB16 cells in relation to the cellsreceiving basal medium. Once again, the pretreatment with JE1:2, JE2:1and JE2:2 compounds reduced said increase and confirms that the JE2:2compound as being the most highly effective. All of the determinationsare made in triplicate.

Later, for the purpose of proving the antioxidant effect of the JE2:2compound, tests were conducted on H202 production by MB16 cells inresponse to recombinant II-18. See FIG. 4, where the treatment with theJE2:2 compound inhibited the production of H202 of cells in basalconditions and completely abrogated (P<0.01) the increase in H2O2 causedby II-18.

Because H₂O₂ released by the hepatic sinusoidal endothelium (HSE)facilitated the adhesion of melanoma cells and the development ofhepatic metastasis, an additional evaluation was acomplished on theeffect of the compounds on tumor-activated HSE cells. As shown in FIG.5, pretreatment of HSE with the JE2:2 compound at a concentration of 2.5μM inhibited endothelial production of H2O2 and significantly (P<0.01)reduced the increase in adhesion of MB16 cells to the HSE activated bysoluble factors of the melanoma (see FIG. 6).

It was also checked whether any of the intermediary compounds in thechemical synthesis of the JE2:2 compound also has the capacity toinhibit tumor cell adhesion. Two of the compounds, called JEM2:2-01 andJEM2:1-O₂ are tested. On the other hand, the preincubation of the MB16cells with 2.5 μM of the JEM2:2-01 compound completely abrogated theincrease in tumor cell adhesion to immobilized VCAM-1 induced by H2O2(see FIG. 7). On the other hand, tumor adhesion to HSE cells treatedwith MC-MB16 in the presence of both of these two compounds wasdetermined. In FIG. 8A, both compounds, JEM2:2-01 and JEM2:1-02,significantly (P<0,01) reduced the proadhesive response induced in theHSE cells by soluble factors from MB16. A greater inhibition wasobserved with JEM2:2-01 compound, indicating that it is the only onewhich completely abrogated the inflammatory response of the endothelium,given that it inhibits the endothelial production of TNF-alpha inducedby the MB-MB16 (see FIG. 8B).

Next, a screening procedure was carried out among the general formula(IV) compounds based on the in vitro tests on H202-treated MB16 celladhesion to immobilized VCAM-1 or to monolayers from primary culturedHSE cells preincubated or not with MC-MB16. All of the compounds tested(YEF02, YEF03, YEF07, YEF05B, YEF07B and YEF05H) significantly (P<0.01)inhibited the increase in adhesion of MB16 cells to immobilized VCAM-1substrate induced by treatment with H2O2 (see FIG. 9) and alsosignificantly (P<0.01) reduced the proadhesive response of HSE tosoluble factors from MB16 cells (see FIG. 10).

Given that the antimetastatic effect of many natural antioxidants, suchas resveratrol, is attributed to the capacity thereof to inhibitcyclooxigenases, an EIA was then conducted to determine PGE2concentration secreted by HSE cells incubated in the presence or absenceof the synthetic compounds and treated or not with VEGF. The compoundstested are those showing a greater inhibitory effect in the previousexperiments, in other words, the JE2:2, YEF07 and YEF05B compounds (seeFIG. 11). All of the compounds significantly inhibited the increase inthe endothelial secretion of PGE2 produced after incubating the HSEcells with 10 ng/ml VEGF for 4 hours. The inhibitory effect was alsoobserved in some cases with the cells cultured under basal conditions.With the concentrations tested, a dose-response was solely observed withthe JE2:2-compound in the presence of the VEGF. Therefore, thesynthesized compounds are somehow regulating the activity of thecyclooxigenases.

In Vivo Test Results:

In following, as study was made as to the in vivo antimetastaticcapacity of the JE2:2 compound by selecting it as the leading compoundof the families studies both for its inhibitory activity of all of theparameters studied in vitro as well as for its being highlywater-soluble. To this end, the MB16 cells were intrasplenicallyinjected in C57BL/6J mice (3×10⁵ viable cells per animal resuspended inendotoxin-free sterile saline solution), 12 days after which thecapacity thereof of development metastasis was determined.

One group of mice (n=7) were administered an intraperitoneal injectionof JE2:2 (2.5 mg/kg) 1 hour prior to tumor cell injection, and the samedose was repeated on days 2, 3, 4, 8, 9, 10 and 11 in following thereto.The group of control mice (n=7) was administered PBS.

As is shown in FIG. 11, the treatment with JE2:2 reduced the meandensity of hepatic metastasis by 87% and the volume occupied bymetastatic tissue by 90%, which meant a statistically significant(P<0,01) antimetastatic effect in comparison to the control miceadministered PBS.

1-25. (canceled)
 26. A nitrogenated trans-stilbene analog compound ofthe following general formula (I):

and salts thereof, wherein: (X) is chosen from imine and pyrrole groups,wherein the pyrrole group may be bonded to the aromatic ring (A) by oneor two of the pyrrole ring carbons; R¹, R², R³, R⁴, R⁵, R⁶, R⁷, R⁸, R⁹and R¹⁰ may be the same or different and are chosen from a hydrogenatom, —OH, and alkoxy groups, wherein at least three of thesesubstituents are chosen from alkoxy-OH and alkoxy groups; and R⁵ isabsent when the pyrrole ring is bonded to the ring (A) by two carbonatoms.
 27. A compound according to claim 26, wherein the compound is ofthe following general formula (II):

and salts thereof.
 28. A compound according to claim 26, wherein (X) isa pyrrole group which may be bonded to the aromatic ring (A) by one ortwo of the pyrrole ring carbons.
 29. A compound according to claim 28,wherein the compound is of the following general formula (III):

and salts thereof, wherein: (Y) is chosen from a hydrogen atom; nitro,amino, linear and branched alkoxycarbonyl, and amide groups; and organicand inorganic quaternary ammonium salts; (W) is chosen from a hydrogenatom, carboxyl, alkoxycarbonyl and aminocarbonyl groups; (Z) is chosenfrom a hydrogen atom, linear and branched alkyl, benzyl, carboxyl,arylmethyl, heteroarylmethyl, O-alkyl(aryl)carbamoyl andN-alkyl(aryl)semicarbazide groups.
 30. A compound according to claim 28,wherein the compound is of the following general formula (IV):

and salts thereof, wherein: (Z) is chosen from a hydrogen atom, linearand branched alkyl, benzyl, carboxyl, arylmethyl, heteroarylmethyl,O-alkyl(aryl)carbamoyl and N-alkyl(aryl)semicarbazide groups; and (U) ischosen from a hydrogen atom and linear and branched alkyl groups.
 31. Acompound according to claim 26, wherein the alkoxy group is a methoxy.32. A compound according to claim 27, wherein the compound is5-((E)-(4-Hydroxyphenylimine) methyl)benzene-1,3-diol.
 33. A compoundaccording to claim 26, wherein (Z) is a hydrogen atom; (W) is chosenfrom —COOCH₃ and —COOH; (Y) is chosen from a hydrogen atom, —NH₂ and—NO₂; R¹, R⁵, R⁶ and R¹⁰ are hydrogen atoms; and R², R³, R⁴, R⁷, R⁸ andR⁹ are the same or different and are chosen from a hydrogen atom, —OCH₃and —OH.
 34. A compound according to claim 33, wherein (Z) and (Y) arehydrogen atoms.
 35. A compound according to claim 33, selected from thefollowing group: 3,5-bis(3,5-Dihydroxyphenyl)-1H-pyrrole-2-methylcarboxylate; 3,5-bis(3,5-Dimethoxyphenyl)-1H-pyrrole-2-methylcarboxylate;5-(3,5-Dihydroxyphenyl)-3-(4-hydroxyphenyl)-1H-pyrrole-2-methylcarboxylate;3-(4-methoxyphenyl)-5-(dimethoxyphenyl)-1H-pyrrole-2-carboxylic acid;5-(3,5-dimethoxyphenyl)-3-(4-methoxyphenyl)-1H-pyrrole-2-methylcarboxylate;5-(3,5-dimethoxyphenyl)-3-(4-methoxyphenyl)-1H-pyrrole-2-methylcarboxylate;5-(3,5-dimethoxyphenyl)-3-(4-methoxyphenyl)-4-nitro-1H-pyrrole-2-carboxylate;4-amino-3,5-bis(3,5-dimethoxyphenyl)-1H-pyrrole-2-methyl carboxylate;5-(3,5-dimethoxyphenyl)-3-(4-methoxyphenyl)-4-nitro-1H-pyrrole-2-methylcarboxylate; 3,5-bis(3,5-dimethoxyphenyl)-4-nitro-1H-pyrrole-2methylcarboxylate; 3,5-bis(3,5-dimethoxyphenyl)-1H-pyrrole-2-carboxylic acid;and 3-(3,5-Dihydroxyphenyl)-5-(4-hydroxyphenyl)-1H-pyrrole-2-methylcarboxylate.
 36. A compound according to claim 26, wherein (Z) is chosenfrom a hydrogen atom and —COOC(CH₃)₃ group; (U) is a hydrogen atom; andR¹, R², R³, R⁴, R⁶, R⁷, R⁸, R⁹ are R¹⁰ the same or different and arechosen from a hydrogen atom, —OCH₃ and —OH.
 37. A compound according toclaim 36, selected from among the following group:2-(2,5-dihydroxyphenyl)-1H-indole-4,6-diol;4,6-Dimethoxy-2-(3,5-dimethoxyphenyl)-1H-indole;4,6-Dimethoxy-2-(2,5-dimethoxyphenyl)-1H-indole-1-tert-butylcarboxylate; 4,6-Dimethoxy-2-(2,5-dimethoxyphenyl)-1H-indole; and4,6-Dimethoxy-2-(3,5-dimethoxyphenyl)-1H-indole-1-tert-butylcarboxylate.38. A method for obtaining the compound according to claim 27,comprising the following steps: i) reacting an aromatic aldehyde with ananiline in the presence or absence of an organic solvent; ii)purification by crystallization in a suitable solvent.
 39. A methodaccording to claim 38, wherein a drying agent is added to the reactionmixture.
 40. A method for obtaining a compound according to claim 29,comprising the following steps: i) reacting an imine, a nitroalkene, ametallic salt and a tertiary organic base; ii) conducting said reactionin conditions selected from a) in the presence of microwave radiationand b) in the presence of an organic solvent, at a temperature rangingfrom −25° C. to +25° C.; iii) obtaining a mixture of 2-alkoxycarbonylpyrrolidines and dissolving said mixture in cyclic ether with anoxidizing agent at a temperature ranging from 60° C. to 250° C.; iv)obtaining a mixture of 2-alkoxycarbonyl-NH-pyrrole and2-alkoxycarbonyl-4-nitro-NH-pyrrole and separating said mixture byfractioned crystallization or chromatography; v) obtaining compound ofgeneral formula (III) by means of chemical transformations of compoundsobtained by iv).
 41. A method for obtaining the compound according toclaim 30, comprising the following steps: i) reacting an amine, analpha-haloketone and a tertiary amine; ii) conducting said reaction inthe presence of microwave radiation at a temperature ranging from 90° C.to 180° C., with a radiation power ranging from 25 W to 200 W, for atime period ranging from 5 minutes to 30 minutes, at a pressure rangingfrom 50 psi to 200 psi. iii) obtaining 2-aryl-1H-indoles from ii); iv)obtaining compounds of general formula (IV) by means of chemicaltransformations of the 2-aryl-1H-indoles from iii).
 42. A methodaccording to claim 41, wherein the temperature at point (ii) ranges from130° C. to 170° C., the radiation power ranges from 80 W to 120 W, in atime ranging from 7 to 15 minutes.
 43. A pharmaceutically-acceptablecomposition comprising at least one compound according to claim
 26. 44.A method of treatment and prophylaxis of carcinogenic and inflammatorydiseases comprising administering a composition according to claim 43.45. A method of treatment according to claim 44 wherein the carcinogenicdisease is hepatic metastasis.
 46. A pharmaceutical compositionaccording to claim 43, in conjunction with at least onepharmaceutically-acceptable vehicle.
 47. A pharmaceutical compositionaccording to claim 43 comprising at least one additional therapeuticallyactive substance.
 48. A pharmaceutical composition according to claim 47wherein the additional therapeutically active substance is quercetin.